Apparatus for controlling movement of specimen, method for controlling movement of specimen, and method for processing specimen

ABSTRACT

An apparatus for controlling the movement of a specimen in order to handle the specimen, e.g., a cell, easily and quickly with high accuracy. A standing wave vibration (V) having at least one node (N 1 , N 2 ) is generated in a vibrator ( 11 ). A specimen in a medium ( 30 ) is captured at a position in the vicinity of the surface of the node (N 1 , N 2 ) and then turned in this position. The capturing position of the specimen (S) is altered through variable control of the vibration mode of the standing wave vibration (V), and the moving route, moving speed, or rotational speed of the specimen (S) is controlled as desired through variable control of the vibration parameters (e.g., frequency, amplitude, intermittent frequency) of the standing wave vibration (V).

TECHNICAL FIELD

[0001] The present invention relates to a technology of controlling themovement of a specimen, and especially a small specimen such as a cell,when it is inspected or subjected to a prescribed treatment.

BACKGROUND ART

[0002] Various manipulations of a small specimen such as a cell aretroublesome operations requiring delicate manipulation with a microtool.One example of such a manipulation is microinjection in a genetictransformation technology. In this genetic engineering technique, a hostcell is pricked with an ultrathin glass tube so to introduce recombinantDNA. Introduction of recombinant DNA into a host cell usingmicroinjection will next be described.

[0003] The host cell as a specimen is introduced to physiological salinein a petri dish. An operator (worker) uses suction to hold the host cellto the tip of a suction pipette, pricks a nucleus of the held host cellwith an ultrathin glass tube, and introduces the recombinant DNA intothe nucleus through the glass tube. While performing this operation, theoperator uses a microscope to observe the procedure and ascertain thatpositions are correct.

[0004] However, because the host cell, which is in a natural orientationin the physiological saline, is suctioned and held without reorientationat the tip of the pipette by the above operation, when the nucleus isnot on the side of the glass tube, in other words when the held specimendoes not have a orientation suitable for manipulation, the operator mustchange the direction of the suction pipette to the specimen repeat theoperation of suctioning and holding the specimen. Thus, the operatormust take considerable time and trouble to obtain a desired orientation.Further, to capture a desired specimen by this manipulation, it isnecessary to move the suction pipette closer to the specimen. In otherwords, this manipulation involves a great deal of effort requiringexperience while observing under a microscope to obtain the desiredorientation of a particular specimen.

DISCLOSURE OF THE INVENTION

[0005] The apparatus for controlling the movement of a specimenaccording to the present invention comprises a vibrator which vibratesin a medium, an oscillation mechanism which causes the vibrator vibrate,and a vibration control mechanism which controls the oscillationmechanism so as to cause the vibrator generate a standing wave vibrationhaving at least one node. By configuring as described above, thespecimen can be reliably captured at the node of the standing wavevibration in the vicinity of the surface of the vibrator and turned inthis position such that the orientation of the specimen can becontrolled as desired. Therefore, the instances wherein the complicatedprocedure described above would be required can be decreased or eveneliminated, and the movement of the specimen can be controlled moreeasily, quicker, and with a higher accuracy.

[0006] The apparatus for controlling the movement of a specimenaccording to the present invention further comprises a vibration modevariable mechanism which can vary a vibration mode of the vibrator.Thus, the position of the node for capturing the specimen can be changedfreely, and the specimen can be more accurately moved to a desiredposition.

[0007] When the vibrator is vibrated continuously, the specimen,especially for certain specimens or media, may become damaged or it maybecome difficult to control the specimens, because the rate of change ofmovement (e.g., a moving speed or rotational speed) of a specimenbecomes high. According to the present invention, the vibration of thevibrator can be switched intermittently to appropriately adjust thevibration energy of the vibrator. Thus, instances of damage to thespecimen are decreased, and the rate of change of movement of thespecimen is adjusted appropriately, so that the movement of the specimencan be controlled more easily, quicker, and with a high accuracy.

[0008] The method for controlling the movement of a specimen accordingto the present invention comprises a step which places the specimen in amedium and a step which causes a vibrator disposed in the medium togenerate a standing wave vibration having at least one node so tocapture the specimen at the node of the standing wave vibration in thevicinity of the surface of the vibrator. Thus, the instances wherein thecomplicated procedure described above would be required can be decreasedor even eliminated, and the movement of the specimen can be controlledmore easily, quicker, and with a higher accuracy.

[0009] The method for manipulating a specimen according to the presentinvention comprises a step which places the specimen in a medium; a stepwhich causes a vibrator disposed in the medium to generate a standingwave vibration having at least one node so to capture the specimen bythe node of the standing wave vibration in the vicinity of the surfaceof the vibrator; and a step which performs a prescribed treatment of thecaptured specimen. Thus, the specimen which is accurately captured in adesired position and/or orientation can be treated, so that theprescribed treatment of the specimen can be performed more easily,quicker, and with a high accuracy.

BRIEF DESCRIPTION OF THE DRAWINGS

[0010]FIG. 1 is an explanatory diagram showing a standing wave vibrationof a vibrator of an apparatus for controlling the movement of specimensaccording to an embodiment of the present invention, and the resultingflow of the mediums and movement of the specimens.

[0011]FIG. 2 is a schematic configuration diagram of an apparatus forcontrolling the movement of a specimen according to the embodiment ofthe present invention.

[0012]FIG. 3 is a schematic configuration diagram of a manipulator ofthe apparatus for controlling the movement of a specimen according tothe embodiment of the present invention.

[0013]FIG. 4 is an explanatory diagram showing a voltage waveformapplied to an oscillation mechanism by an oscillation mechanism drivecircuit of the apparatus for controlling the movement of a specimenaccording to the embodiment of the present invention.

[0014]FIG. 5 is a flowchart showing a method for treating a specimenusing the apparatus for controlling the movement of a specimen accordingto the embodiment of the present invention.

[0015]FIG. 6 is an explanatory diagram showing a method for removing thecontents of a cell by means of the apparatus for controlling themovement of a specimen according to the embodiment of the presentinvention.

BEST MODE FOR CARRYING OUT THE INVENTION

[0016] An embodiment which applies the apparatus for controlling themovement of a specimen according to the present invention to the controlof the movement of a cell will be described with reference to thedrawings. FIG. 1 shows a standing wave vibration of a vibrator and theresulting a flow of a medium and the movement of specimens, FIG. 2 showsa schematic configuration of the apparatus for controlling the movementof the specimen, FIG. 3 shows a schematic configuration of amanipulator, and FIG. 4 shows a voltage waveform to be applied to anoscillation mechanism.

[0017] First, a principle of the control of the position of a specimenin this embodiment will be described. FIG. 1 shows a state that avibrator 11 (e.g., a probe section 11 b) is oscillated in a medium 30,and a standing wave vibration V, e.g., vertical vibration, is generatedat a tip of the vibrator 11 which is immersed in the medium 30. It wasfound through research conducted by the present inventors that when thevibrator 11 vibrates with the standing wave vibration V having at leastone node (N1, N2), the following phenomenon is caused.

[0018] (I) Specimens S in the medium 30 move from the periphery of thevibrator 11 toward the nodes (N1, N2) of the standing wave vibration Vand remain at positions in the vicinity of the surface of the nodes (N1,N2).

[0019] Utilizing the above characteristic, in this embodiment thevibrator 11 generates a standing wave vibration V having at least onenode (N1, N2) to capture the specimens S in the medium 30, in thepositions in the vicinity of the surface of the node (N1, N2) Here,positions where the specimens S are captured can be controlled freelythrough variable control of the vibration mode of the standing wavevibration V, and the specimens S can be controlled to a desired movementroute or moving speed through variable control of vibration parameters(e.g., frequency, amplitude, intermittent frequency) of the standingwave vibration V.

[0020] The present inventors also found that the following phenomenon isobserved when the vibrator 11 is vibrating at the standing wavevibration V having at least one node (N1, N2) as described above.

[0021] (II) The specimens S which are caught in the positions in thevicinity of the surface of the nodes (N1, N2) of the standing wavevibration V are rotated in these positions by the standing wavevibration V.

[0022] Utilizing the above characteristic, in the present embodiment thespecimens S, which are captured in the positions in the vicinity of thesurface of the nodes (N1, N2), are rotated by the standing wavevibration V. The specimens S can then be controlled to rotate in adesired rotating direction or at a desired rotational speed usingvariable control of the vibration parameters (e.g., frequency,amplitude, intermittent frequency) of the standing wave vibration V.Thus, the movement and/or orientation of the specimens S can becontrolled as desired. As shown in FIG. 1, the specimens S at theadjacent nodes (N1, N2) rotate in different directions.

[0023] The specimen movement control apparatus 1, which freely controlsthe movement of the specimens S according to the aforementionedprinciple has a manipulator 10 and a control unit 20 as shown in FIG. 2.The manipulator 10 has a vibrator 11, an oscillation mechanism 12 foroscillating the vibrator 11, and a support section 13 for supporting thevibrator 11 as shown in FIG. 3. The vibrator 11 in turn has a vibratorbase 11 a, e.g., a metal tube, having the oscillation mechanism 12, aprobe section 11 b, e.g., a glass tube, and a tip of which is immersedin the medium 30, all connected by a joint section 11 c, e.g., a rodmember.

[0024] The probe section 11 b is detachably fitted to the vibrator base11 a at the joint section 11 c. A plurality of probe sections 11 b canbe exchangeably fitted to the joint section 11 c. The vibration mode ofthe vibrator 11 is variable depending on a shape or a material of thevibrator 11. Therefore, when the vibrator 11 is configured to have adetachable component (e.g., the probe section 11 b) with a differentshape or material, the movement of the specimen S can be controlled byan appropriate vibration mode according to the characteristic (e.g., asize, mass or the like of the specimen S) of the specimen S or thecharacteristic (e.g., a viscosity or the like) of the medium 30.Specifically, the joint section 11 c and the probe section 11 bcorrespond to a vibration mode variable mechanism which changes thevibration mode of the vibrator 11 by exchanging a component (e.g., theprobe section 11 b) of the vibrator 11. The vibrator 11 of thisembodiment has another component (e.g., the probe section 11 b) which isdetachable to the component, e.g., the vibrator base 11 a, having theoscillation mechanism 12. Thus, it is easy to change the shape of thevibrator 11 and the vibration mode of the vibrator 11 while keeping thestate of the oscillation mechanism 12 fitted to the vibrator 11.

[0025] The oscillation mechanism 12 is fixed to the vibrator 11, e.g.,the vibrator base 11 a. In this embodiment, the oscillation mechanism 12is, for example, a cylindrical piezoelectric ceramic element which has asimilar cylindrical detection element 12 b fitted to surround acylindrical oscillator 12 a, and it is tightly fitted to surround theexterior of the vibrator base 11 a which is a metal tube. The oscillator12 a and the detection element 12 b are connected to an oscillationmechanism drive circuit 22.

[0026] Next, the control unit 20 will be described. The control unit 20has a control section 21, e.g., a CPU, for controlling the entireapparatus 1, the oscillation mechanism drive circuit 22, e.g., a pulsegenerator, for driving the oscillation mechanism 12, a memory section23, e.g., a RAM/ROM, an input section 24 (e.g., a keyboard 24 a, a speedadjustment knob 24 b, a movement/stop changeover switch 24 c), and anoutput section 25, e.g., a display.

[0027] The control section 21 controls vibration of the oscillationmechanism 12 by controlling the oscillation mechanism drive circuit 22,to cause the vibrator 11 generate a standing wave vibration having atleast one node. Specifically, the control section 21 corresponds to thevibration control mechanism. More specifically, the control section 21controls the vibration of the oscillation mechanism 12 by controlling apulse voltage (e.g., a pulse voltage shown in FIG. 4) which is appliedfrom the oscillation mechanism drive circuit 22 (e.g., a pulsegenerator) to the oscillation mechanism 12.

[0028] By employing the configuration described above, the controlsection 21 is able to change a frequency f (=1/(2πT), T: cycle) of theapplied voltage. This frequency f substantially corresponds to thevibration frequency of the vibrator 11, so that the vibration mode ofthe vibrator 11 is changed. In other words, the control section 21corresponds to the vibration mode variable mechanism. The higher thefrequency f, the greater will be the number of nodes. The controlsection 21 adjusts the amplitude A of the applied voltage so to adjust amoving speed (e.g., a moving speed or a rotational speed) of thespecimen S in the medium 30. The higher the amplitude A of the appliedvoltage (namely, the vibration of the vibrator 11 has higher amplitude),the faster will be the moving speed of the specimen S. Additionally, thecontrol section 21 periodically switches the applied voltage ON or OFFwith pulsed intermittency (i.e., the vibration of the vibrator 11 isintermittently generated) to control a frequency fp (=1/(2πTp), Tp: onecycle of intermittence/stop, hereinafter the frequency fp will be calledas an intermittent frequency fp) for switching the intermittenceperiodically and a ratio between a vibrating period and a vibrationstopping period (hereinafter called as the intermittent duty ratio). Thehigher the intermittent frequency fp, the faster is the moving speed orthe rotational speed. Additionally, the moving speed of the specimen Sbecomes faster as the vibrating period becomes longer (as theintermittent duty ratio is higher). In other words, when the appliedvoltage to the oscillation mechanism 12 has a waveform as shown in FIG.4(b), the specimen S has a moving speed faster than that shown in FIG.4(a).

[0029] The memory section 23 stores the vibration parameters (thevibrator 11 [e.g., the probe section 11 b], vibration frequency,amplitude or intermittent frequency) for each treating condition (e.g.,a type, size and mass of the specimen S to be treated or a medium fortreating) of the specimen S. Thus, the control section 21 readsappropriate vibration parameters according to the treating conditionsand can quickly and accurately control the vibration of the vibrator 11and, therefore, the movement of the specimen S.

[0030] The input section 24, for example the keyboard 24 a, inputs andsets the treating conditions, the vibration parameters, and the like.According to the input through the input section 24, the control section21 controls each section of the apparatus 1. The speed adjustment knob24 b inputs an instruction regarding a moving speed or a rotationalspeed of the specimen S to the control section 21. More specifically,the control section 21 controls, for example, an intermittent frequencyso as to change the moving speed or the rotational speed of the specimenS according to the amount a the knob 24 b is turned from its neutralposition NP. The memory section 23 stores the intermittent frequency atthe neutral position NP and, when the knob 24 b is turned in a clockwisedirection from the neutral position NP, the control section 21 increasesthe intermittent frequency to higher a level higher so as to increasethe moving speed of the specimen S, and when the knob 24 b is turned ina counterclockwise direction, the control section 21 lowers theintermittent frequency so as to slow the moving speed of the specimen S.The movement/stop changeover switch 24 c switches movement/stop of themovement or rotation of the specimen S by manual operation for example.More specifically, this switch 24 c may be configured as a push button,for example,and the vibrator 11 may vibrate when the button is depressedand cease vibrating and when the button is no longer pushed. In otherwords, the control section 21 switches on and off oscillation of thevibrator 11 according to the input operation of the switch 24 c.Therefore, the operator can operate the switch 24 c to stop the movementof the specimen S in a desired position or orientation. The outputsection 25, for example a display, outputs, for example displays, thetreating condition or the vibration parameters.

[0031] Next, a specimen treating procedure using the specimen movementcontrol apparatus according to this embodiment will be described. FIG. 5is a flowchart showing the specimen treating procedure.

[0032] First, the specimen movement control apparatus 1 is set(apparatus setting preparation step S1). In this step S1, the vibrator11 is fitted to the specimen movement control apparatus 1, and the probesection 11 b is fitted to the joint section 11 c. As the vibrator 11 andthe probe section 11 b fitted here, those which generate an appropriatevibration mode depending on the specimen S and the medium 30 areselected.

[0033] Then, the medium 30 is poured into a vessel, and the specimen Sis placed in the medium 30 (step S2 for placing the specimen into themedium).

[0034] The display 25 can display the past treatment conditions orvibration parameters stored in the memory section 23. For example, itmay judged by the operator from the displayed results whether the storedcondition setting is used or not (use condition selection step S3),and,when the stored condition setting is not used, vibration parameters areinput through the input section 24, e.g., the keyboard 24 a (vibrationparameter input step S4). When the stored condition setting is used, thecontrol section 21 obtains the vibration parameters of the conditionsetting from the memory section 23 according to the input about it fromthe input section 24, e.g., the keyboard 24 a.

[0035] Next, the control section 21 begins oscillation of the vibrator11 (vibration start step S5). For example, it is judged by observingthrough a microscope whether the specimen S is caught in a desiredposition and/or orientation (step S6 for judging capture in a desiredposition/orientation), and when the specimen S is not caught in adesired position/orientation, an instruction is input from the inputsection 24, e.g., the keyboard 24 a, the speed adjustment knob 24 b, orthe movement/stop changeover switch 24 c, and the position and/ororientation of the specimen S is adjusted (specimen position/orientationadjustment step S7). More specifically, the moving speed or therotational speed of the specimen S is adjusted by operating the speedadjustment knob 24 b for example, and the movement/stop of the specimenS is switched by operating the movement/stop changeover switch 24 c toadjust the specimen S to a desired position and/or orientation. Thecapturing position can be moved along the surface of the vibrator 11 ora flow of the medium 30 around the vibrator 11 can be changed byinputting a change instruction for the vibration frequency of thevibrator 11 through the keyboard 24 a, for example.

[0036] After capturing in a desired position and/or orientation iscompleted, the treatment of the specimen S, e.g., microinjection intothe cell nucleus, is performed. In this embodiment, because the specimenS can be captured in a position and a orientation suitable for itstreatment, the treatment can be performed easily and quickly with highaccuracy. For example, when an egg cell is used as the specimen S in themicroinjection, the vibration of the vibrator 11 may be controlled toadjust the orientation of the egg cell so to direct the nucleus of theegg sell toward the injector.

[0037] Before the power of the specimen movement control apparatus 1 isturned off, the control section 21 stores the vibration parameters usedfor controlling the position and/or orientation of the specimen in thememory section 23 (control condition storing step S9). Thus, when asubsequent treatment is performed under the same condition (e.g., theused specimen or medium 30), the position and/or orientation of thespecimen S can be controlled still more quickly and more accuratelyusing the stored data.

[0038] Next, a method for removing the contents of a cell as thespecimen by the specimen movement control apparatus 1 will be described.FIG. 6 is an explanatory diagram showing a state of the cell treated bythis method.

[0039] First, the above-described steps S1 to S7 are performed tocapture a specimen having contents, e.g., a cell 50, in a desiredposition and orientation (FIG. 6(a)). Then, a needle 60 is inserted intoa shell, e.g., cell membrane 51, of the specimen to form a hole 52 (cellmembrane opening step; FIG. 6(b)). According to an instruction inputfrom the input section 21, the cell 50 is rotated (cell rotation step;FIG. 6(c)). When the cell 50 is rotated, the contents 53 of the cell 50are discharged out of the cell 50 by centrifugal force to the medium 30through the hole 52. By the above method, the shell, e.g., the cellmembrane 51, having the contents 53 removed can be obtained easily andquickly (FIG. 6(d)). By appropriately setting the rotational speed ofthe cell in its rotation step depending on the treating conditions(e.g., a type, size and mass of the cell 50, a size of its contents[e.g., a nucleus] or a viscosity of the contents), the contents 53 canbe removed more quickly and more accurately.

[0040] The present invention is not limited to the above-describedembodiment. For example, a mechanism which has a member in contact withthe vibrator and can change its contact position may be disposed as thevibration mode variable mechanism. By configuring as described above,the vibration mode of the vibrator can also be controlled variably. Thespecimen movement control apparatus may also be provided with an imagepickup section for imaging of the specimen and a position/orientationjudgment section which judges whether the specimen is captured in adesired position or orientation according to the image taken by theimage pickup section, and the control of the specimen to a desiredposition/orientation may be performed automatically according to thedetermined position/orientation.

INDUSTRIAL APPLICABILITY

[0041] As described above, according to the specimen movement controlapparatus, method, and the specimen treating method according to thepresent invention, the movement of a specimen in the field ofbiotechnology can be controlled freely by making the vibrator disposedin the medium generate a standing wave vibration having at least onenode, so that, for example, a very small specimen such as a cell can betreated easily, quickly, and with a high accuracy.

1. An apparatus for controlling the movement of a specimen, comprising:a vibrator which vibrates in a medium; an oscillation mechanism whichcauses the vibrator to vibrate; and a vibration control mechanism whichcontrols the oscillation mechanism to make the vibrator generate astanding wave vibration having at least one node.
 2. The apparatus forcontrolling the movement of a specimen according to claim 1, furthercomprising a vibration mode variation mechanism which can change thevibration mode of the vibrator.
 3. The apparatus for controlling themovement of a specimen according to claim 1, wherein the vibrationcontrol mechanism causes the vibrator to vibrate intermittently.
 4. Amethod for controlling the movement of a specimen, comprising: a step ofplacing the specimen in a medium; and a step of causing a vibratordisposed in the medium to generate a standing wave vibration having atleast one node so as to capture the specimen by the node of the standingwave vibration in the vicinity of the surface of the vibrator.
 5. Amethod for manipulating a specimen, comprising: a step of placing thespecimen in a medium; a step of causing a vibrator disposed in themedium to generate a standing wave vibration having at least one node soas to capture the specimen at the node of the standing wave vibration inthe vicinity of the surface of the vibrator; and a step of performing aprescribed treatment of the captured specimen.